The together cell culture model used in this project is an embryonic mouse brain co culture that includes neurons, astrocytes and microglia, in order to reflect the cell population in normal adult mouse cortex. In control conditions without amy loid stress, no inflammatory reactive glia were observed, excluding any trauma during cell preparation. The major aim with this model was to be close to physiolo gic conditions and to recreate in vitro the essential neu ron glia environment to explore the effects of the inflammatory process on neurons. Currently, indepen dent cultures of microglia or astrocytes with or without neurons are widely used as models of inflammation in brain. However, it seemed essential to maintain these three cellular actors together in our experimental condi tions, considering the multiple interactions between neurons and glia, in particular in inflammatory condi tions.
This model is produced from embryonic tissue, and one must therefore remain cautious about its use because, as we know, Inhibitors,Modulators,Libraries the maturity of the regulatory and compensation processes Inhibitors,Modulators,Libraries is not complete. The cells may be more or less vulnerable to the toxicity of amy loid peptide compared with adult cells. Their tolerance system has not yet been sufficiently explored. In addi tion, the concentration of exogenous amyloid peptide added in cultures, although identical to that used in many published studies, is far greater to that found in brains of patients with Alzheimers disease. However, it is known that levels of both Abx 40 and Abx 42 increase very early in the disease process, and in the frontal cortex these increases occurr in the absence of significant neurofibrillary pathology.
These levels increase systematically with severity of cognitive decline contrary to Ab burden as assessed only in neuritic plaques. For this mixed co culture Inhibitors,Modulators,Libraries model, we have shown that the PKR inhibitor at a concentration of 1 uM, as the morphology of microglia. In Ab conditions, many of the neurons showed signs of neuritic damage with bead ing and fragmentation, according Inhibitors,Modulators,Libraries to other studies, and formation of pleiomorphic microglia was observed with ramified microglia and features of chroni cally activated Inhibitors,Modulators,Libraries microglial cells represented by a markedly elongated cells named rod microglia. In brains of patients with AD, activated rod and ramified microglia are observed, ramified microglia are in contact with amyloid fibrils and rod microglia are found predomi cell assay nantly at the edge of senile plaques. For astro cytes, morphological modifications were very limited with thinner extensions. This mixed co culture model of previously used on neuroblastoma cell line, induces a great alteration, leading us to use a lower concentra tion of 210 nM corresponding to the IC50.